Meeting Abstract

S11-1  Thursday, Jan. 7 08:00  Alternative approaches to building transcriptomes can impact differential gene expression analyses in crustaceans exposed to high CO2 and low O2. JOHNSON, Jillian G.*; BURNETT, Louis E.; BURNETT, Karen G.; College of Charleston

Aquatic crustaceans can experience hypoxia (low O2) and hypercapnic hypoxia (high CO2/low O2) both in the natural estuaries and in aquaculture ponds. RNA-Seq data were collected from the hepatopancreas of aquacultured Pacific whiteleg shrimp Litopenaeus vannamei (n=6/treatment x time) exposed to short term (4 h) or prolonged (24 h) hypoxia, hypercapnic hypoxia, or air-saturated water (normoxia) to explore the global transcriptomic response and specific involvement of the suite of hemocyanin (Hc) subunits that make up the higher order O2-transport protein. Assembly using the robust de novo assembler, Trinity, produced a high-quality stranded transcriptome (assembly size, 52,190 contigs; mean length, 870 bp; N50 1680 bp; RMBT%, 96.03%); however, the highly expressed Hc gene family (hemocyanins, prophenoloxidases, cryptocyanins) experienced high rates of assembly misalignments, hybrids, and partial fragments. Manual curation of the Hc subunits revealed 1) the large Hc subunit has more sequence diversity than previously identified (HcL1-3 isoforms) and 2) penaeid shrimp express a β-type Hc subunit (HcB). Transcriptome assemblies of wild caught L. vannamei and Atlantic brown shrimp Farfantepenaeus aztecus demonstrated the same Hc transcript assembly misalignments and hybrid transcripts, but highly expressed transcripts from single copy gene loci assembled into full-length contigs. High occurrence of single nucleotide polymorphisms (SNPs) coupled with high sequencing error rates for overrepresented sequences, like the hemocyanins, have a profound affect on assembly accuracy and thus downstream differential expression analyses. (NSF IOS-1147008)